Depurination, the loss of a purine base from DNA, has been shown to be highly mutagenic in a forward mutational assay using M131ac DNA. DNA sequence analysis of 213 mutants indicates that this DNA damage primarily results in base substitution mutations, while only a small increase in frameshift mutagenesis is observed. The mutagenicity is predominantly observed in the presence of error prone DNA synthesis and exhibits a strong specificity for insertion of dAMP opposite the apurinic site, resulting primarily in transversions. These data in a forward mutational system provide insight into the mechanisms used by a cell to replicate DNA containing non-coding lesions.